Master replication origins at the heart of the organisation and fragility

of the human genome

Alain ArneodoLaboratoire Joliot-Curie / Laboratoire de Physique

Ecole Normale Supérieure de Lyon46 allée d’Italie, 69364 Lyon Cedex 07, France

Alain.Arneodo@ens-lyon.fr

Benjamin Audit

Edward-Benedict Brodie of Brodie

Samuel Nicolay ENS de Lyon, France

Philippe Saint-Jean

Cédric Vaillant

Lamia Zaghloul

Yves d’Aubenton-Carafa

Maxime Huvet

Claude Thermes CGM, Gif-sur-Yvette, France

Marie Touchon

DeoxyriboNucleic Acid

A

GC

T

G C

T A

: :

• Double helix macromolecule

• Each strand consists of an oriented sequence of four possible nucleotides:Adenine, Thymine, Guanine & Cytosine

• Complementary strands:[A]=[T] & [G]=[C] over the sum of both strands

Sequencing projects result in 4 letter texts :

NET RESULT : EACH DNA MOLECULE HAS BEEN

PACKAGED INTO A MITOTIC CHROMOSOME THAT IS 50.000x SHORTER THAN ITS EXTENDED LENGTH

HIERARCHICAL STRUCTURE OF EUCARYOTIC DNA

LARGE SCALE REPRESENTATION OF GENOMIC SEQUENCES

Chromosome 22 (Human)

Nicolay, Phys. Rev. Lett. (2004)

Strand Compositional AsymmetryStrand Compositional Asymmetry

Transcription

Replication

Opening of the double helix with a different environment for each strand => asymmetrical process

[A] = [T] & [G] = [C]

in each strand

Deviations from this property estimated by the compositional skews

Compositional skew due to local biases in a strand in the course of biological mechanisms

Symmetrical properties of the strands: ‘‘Parity Rule type 2’’

S = [C] – [G]

[C] + [G]CG

S =[A] – [T]

[A] + [T]AT

x 106 pb

SSG

CG

C

G > CG < C

5’5’ 3’3’

Bacillus subtilisBacillus subtilis

leading strandleading strandlagging strand

ORIORI

TERTERTERTER

S < 0 S > 0

Replication bias in bacteria:An upward jump at the origin

ORI

TER

ORIORI ORIORITERTER

x 106 pb

SSG

CG

C

lagging strandleading strand

G < CG > C

5’5’ 3’3’

Bacillus subtilisBacillus subtilis

S > 0 S < 0

Replication bias in bacteria:A downward jump at the terminus

ORI

TER

Skew profiles around human replication origins

upward jumps : ∆S ~ 24%

Can these profiles be explained by transcription only ?

Do intergenic regions show upward transitions of the skew S ?

Brodie of Brodie, Phys. Rev. Lett. (2005)Touchon, PNAS (2005)

Conservation of profiles in mammalian genomes

human

mouse

rat

dog

Touchon, PNAS (2005)

Multi-scale detection of jumps in skew profiles using the wavelet transform

Wavelet transform of the skew profileHuman chromosome 6

S=

SG

C+

ST

A

5’ 3’

ORIORI

S

0

ORIORI

d d

Transcription biasH

uman

Statistics of upward and downward jumps at scale 10 kbp

∆S>0.1, ∆n<2kbp: 36% (7228/20023) significantly biased human genes likely expressed in germ line cells.

T.I. Lee et al., Cell 125 (2006): 32% of human ES cell genes bound to Pol II

Nicolay, Phys. Rev. E (2007)

Jumps in the skew profiles at scales a > 200kbp

S=

SG

C+

ST

A

Statistics of upward and downward jumps in Human skew profiles

« Factory roof » profiles

S (%)

TOP1

MCM4

S (%)

Skew profiles along 9Mb Human contigsBrodie of Brodie, Phys. Rev. Lett. (2005)Touchon, PNAS (2005)

In germ-line cells, termination sites are regularly dispersed between adjacent origins

Model of ReplicationWell positionned replication origins,

separated by diffuse terminiBrodie of Brodie, Phys. Rev. Lett. (2005)Touchon, PNAS (2005)

Replication domain detection using an adapted analyzing wavelet

Audit, Phys. Rev. Lett. (2007)Huvet, Genome Res. (2007)

ASYMMETRY OF HUMAN GENOME

factory roofs = 36 %

factory roofs < 1 %

size (kbp)

oriorioriori

oriorioriori

distance to putative origins (Mbp)

oricandidatescandidates

oricandidatescandidates

The detected jumps are replicated earlier than surrounding regions

Av

era

ge

tim

ing

re

pli

ca

tio

n p

rofi

le

Woodfine et al., Cell Cycle (2005)

Human chromosome 6

Audit, Phys. Rev. Lett. (2007)Huvet, Genome Res. (2007)

The predicted origins replicate earlier than their surrounding regions

: Average timing ratio profile around the 83 putative chr6 origins : 20 origins with well defined local maxima : 10 late replicating originsTiming ratio data from Woodfine et al., Cell Cycle (2005)

Audit, Phys. Rev. Lett. (2007)Huvet, Genome Res. (2007)

Disentangling replication and transcription contributions to skew profiles

Audit, Phys. Rev. Lett. (2007)

QuickTimeﾪ et undﾪcompresseur TIFF (non compressﾪ)

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QuickTimeﾪ et undﾪcompresseur TIFF (non compressﾪ)

sont requis pour visionner cette image.

Abundance of small nascent replication strands

Replication timing profile

Genome-wide high-resolution (kbp) experimental verification of putative replication origins using

massive sequencing technology

Olivier Hyrien’s group - ENS - Paris

Genome organisation around putative replication origins

Huvet, Genome Res. (2007)

Gene distribution around predicted replication origins

dd

kbp

NN

5’ 3’

ORIORI

S

0

ORIORI

d d

s

kew

pro

file

Distribution of gene length around predicted replication origins

Mbp

putativeputativeorigins

putativeputativeorigins

kbp

distance from replication originsdistance from replication origins

LL

Genome organisation around putative replication origins

transcribed regions around putative origins (± 20 kb) :

• 55 % are oriented as replication fork progression

• 5% are in opposed orientation

• 35% are intergenic

Gene expression around predicted replication origins

distance from replication origins (kb)

N

ESTsESTs

Bre

adth

of

exp

ress

ion

QuickTimeﾪ and aTIFF (Uncompressed) decompressor

are needed to see this picture.

Model for the large-scale functional organisation of the human genome

Huvet, Genome Res. (2007)

Genomic DNA codes for open chromatin around “master” replication origins

Human chromosome 6

Audit, Nucleic Acids Res. (2009)

Genomic DNA codes for open chromatin around “master” replication origins

Dnase HS sites coverageNFR densityCpG o/e

• R+ gene coverage

Genomic DNA codes for open chromatin around “master” replication origins

Fragility of the human genome around “master” replication origins

Cancer gene Evolutionarybreakpoints

Audit, Nucleic Acids Res. (2009)Lemaitre, BMC Genomics (2009)

Genomic DNA codes for fragile open chromatin around “master” replication origins

Dnase HS sites coverageNFR densityCpG o/e

• R+ gene coverage

Dnase HS sites coverageNFR densityCpG o/e

• R+ gene coverage

Genomic DNA codes for fragile open chromatin around “master” replication origins

Dnase HS sites coverageNFR densityCpG o/e

• R+ gene coverage

Genomic DNA codes for fragile open chromatin around “master” replication origins

A cascading domino model for the spatio-temporal replication program in mammalian cells

QuickTime et unﾪdﾪcompresseur TIFF (non compressﾪ)

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DNA replication timing data corroborate in silico human replication origin predictions

B. AUDIT, S. NICOLAY, M. HUVET, M. TOUCHON, Y. D'AUBENTON-CARAFA, C.THERMES & A. ARNEODO. Phys. Rev. Lett. 99, 248102 (2007).

Human gene organization driven by the coordination of replication and transcription

M. HUVET, S. NICOLAY, M. TOUCHON, B. AUDIT, Y. D’AUBENTON-CARAFA, A. ARNEODO.& C. THERMES. Genome Res. 17, 1278-1285 (2007).

Bifractality of human DNA strand-asymmetry profiles results from transcription.

S. NICOLAY, E.B. BRODIE OF BRODIE, M. TOUCHON, B. AUDIT, Y. D’AUBENTON-CARAFA, C. THERMES & A. ARNEODO. Phys. Rev. E 75, 032902 (2007).

DNA in chromatin: from genome-wide sequence analysis to the modeling of replication in mammals.

A. ARNEODO, Y. D’AUBENTON-CARAFA, B. AUDIT, E.B. BRODIE OF BRODIE, S. NICOLAY, P.ST-JEAN, C. THERMES, M. TOUCHON, & C. VAILLANT. Adv. Chem. Phys. 135, 203 (2007).

Replication-associated strand asymmetries in mammalian genome : towards detection of replication origins

M. TOUCHON, S. NICOLAY, B. AUDIT, E.B. BRODIE OF BRODIE, Y. D’AUBENTON-CARAFA, A. ARNEODO & C. THERMES, Proc. Natl. Acad. Sci.USA 102, 9836-9841 (2005)

From DNA sequence analysis to modeling replication in the Human genomeE.B. BRODIE OF BRODIE, S. NICOLAY, M. TOUCHON, B. AUDIT, Y. D’AUBENTON-CARAFA,

C. THERMES & A. ARNEODO, Phys. Rev. Lett. 94, 248103 (2005)

Transcription-coupled and splicing-coupled strand asymmetries in eukaryotic genomes.

M. TOUCHON, A. ARNEODO, Y. D’AUBENTON-CARAFA & C. THERMES, Nucleic Acids Res. 32, 4969 (2004)

Low Frequency rhythms in human DNA sequences : a key to the organization of gene location and orientation?

S. NICOLAY, F. ARGOUL, M. TOUCHON, Y. D’AUBENTON-CARAFA, C. THERMES & A. ARNEODO, Phys. Rev. Lett. 93, 108101 (2004)

Transcription-coupled TA and GC strand asymmetries in the human genome.M. TOUCHON, S. NICOLAY, A. ARNEODO, Y. D’AUBENTON-CARAFA & C. THERMES,

FEBS Letters 555, 579 (2003)

REFERENCES

Eukaryotic NucleusNative Chromatin fiber

Laboratoire Joliot-Curie

Multi-scale detection of jumps in human skew profile

identification of transitions

Detection of 2415 upward jumps and of a similar number of downward jumps

position of transitions

Scale analysis : 200 kbp

Wavelet based analysis

Sca

le (

kb

)S

cale

(k

b)

SS

Human chromosome 12

2.0

1.5

1.0

n

Re

plic

atio

n tim

ing

Ske

w p

rofil

e

Replication and genome organization

Replication bias in Bacillus subtilis

Sense genes, anti-sense genes

Transcription bias

Replication bias in bacteria

TERORI ORI

Hum

an

Replication and genome organization

High-resolution mapping of Open ChromatinBoyle et al., Cell 132, 311 (2008)

Human chromosome 6

High-resolution mapping of Open ChromatinBoyle et al., Cell 132, 311 (2008)

Human chromosome 6

The sequence is highly predictive of open chromatin around putative replication origins

The sequence is highly predictive of open chromatin around putative replication origins